Ductal epithelial cells and/or centroacinar cells are believed to harbor progenitor cells and give rise to the common pancreatic adenocarcinoma. While there are experimental models to suggest that there is significant capacity for regeneration within the adult pancreatic tissue, considerable debate exists over whether these events represent true examples of "transdifferentiation" as opposed to selective expansion of undifferentiated progenitors in ductal cells or centroacinar cells. Furthermore, the molecular basis for pancreatic ductal branching morphogenesis during exocrine development and epithelial carcinogenesis has yet to be elucidated, which might provide new clues as to the events that trigger ductal differentiation, potential transdifferentiation into other cell types, and malignant transformation. The PDX-1 homeodomain transcription factor is of critical importance in endocrine cell lineage specification;however, the molecular regulation of exocrine (specifically ductal cell lineage) remains elusive. We describe in vitro and in vivo evidence that PDX-1 plays a crucial role in the regulation of ductal morphogenesis, using cytokeratin 19 as a surrogate of ductal gene expression. The overarching hypothesis of this proposal is that PDX-1 modulates pancreatic ductal branching morphogenesis, and this goes awry during malignant transformation. Using innovative three-dimensional cell culture systems (spheroid cysts) and Pdx-1 C-terminal mutant knock-in mice, this hypothesis will be pursued by the following interrelated Specific Aims: (1) To determine the molecular mechanisms underlying pancreatic ductal formation, differentiation and migration, termed branching morphogenesis or tubulogenesis, in a three-dimensional cell culture system;(2) To determine the in vivo role of PDX-1 in pancreatic ductal branching morphogenesis;and (3) To characterize terminal intercalated duct-acinar junctional cells and to characterize known factors regulating centroacinar cell proliferation, differentiation and gene expression. The insights gained from these complementary approaches will provide new insights into how pancreatic ducts develop, how they migrate and compartmentalize, and how they undergo malignant transformation. These studies will provide a platform for new molecular diagnostics and targeted therapeutics.